Mixed Genotype Research Articles

Unveiling the genotypic diversity of Candida albicans and Candida dubliniensis in the oral cavities of drug abusers in Ahvaz, Iran

Candida albicans is a diploid yeast that, under certain conditions, can cause oral or oropharyngeal infections, particularly in immunocompromised individuals. Recent molecular investigations have classified genotypes A, B, and C for Candida albicans, along with genotype D for Candida dubliniensis. This study aimed to identify the different genotypes of the C. albicans complex in drug abusers in Iran. Oral swabs were collected from drug abusers and cultured on CHROMagar Candida. A 21-plex PCR method was employed for the detection of isolates, and the Candida 25S rDNA gene was amplified using primer pairs CA-INT-L and CA-INT-R for ABC genotyping of C. albicans. Out of the 245 substance abusers screened, 151 individuals (61.63 %) were found to harbor the C. albicans complex. The most common genotype among patients was genotype D (39.1 %), followed by genotype A (31.12 %), genotype B (9.93 %), and genotype C (5.29 %). Additionally, 14.56 % of patients had a mixed genotype. Notably, significant differences in genotype distribution were observed in relation to age, underlying diseases, and marital status (P < 0.05). This study highlights the significance of molecular genotyping in understanding the epidemiology of C. albicans and C. dubliniensis in at-risk groups.

Hepatitis B viral infection and its common genotype circulation: A molecular approach

Hepatitis B virus (HBV) is the causes of liver infection and associated with cirrhosis, carcinoma, and fibrosis of liver. In this research study, we conducted an observational study to evaluate the distribution of HBV infection and its genotypes in selected cities of Khyber Pakhtunkhwa, Pakistan. A total of 426 samples were collected from selected cities of Khyber Pakhtunkhwa. Mardan (n = 163), Dir (Lower) (n = 116), Malakand (n = 53), Miran Shah (n = 29), Dera Ismail Khan (n = 28), Swat (n = 22), and Hangu (n = 15). For molecular identification of hepatitis B virus infection, the ICT (Immunochromatographic techniques) kits were used for the Screening of Hepatitis B virus infection among general population. The positive samples of ICT test were further subjected to Real-time Polymerase chain reaction (PCR), of which 282 (66.2 %) were males, and 144 (33.8 %) were females. Out of 426 ICT kit positive samples, 218 (51.18 %) were found PCR positive. The maximum number of positive cases, 157 (36.86 %), was found between 15 and 34 years. Sixty samples were further processed for HBV genotyping. A mixed genotype was detected in 29 (48.3 %) individuals, followed by genotype D in 23 (38.3 %) and untypable in 8 (13.3 %). The positivity rate of HBV was recorded high among males compared to females. Most patients who reported positive for HBV were co-infected with two genotypes. This research will help to provide background information to government and health authorities to raise public awareness and improve the capacity of the healthcare delivery system for hepatitis B virus infection.

High efficacy and safety of direct-acting antivirals for the treatment of chronic hepatitis C: A cohort study conducted in Vietnam.

Direct-acting antivirals (DAAs) have revolutionized hepatitis C virus (HCV) treatment through their high cure rates and improved safety profiles. We aimed to evaluate the efficacy and safety, and identify the optimal combination, of DAAs for the treatment of chronic HCV. A retrospective study was conducted of 613 patients with chronic HCV who were treated with DAAs. Demographic, HCV genotype, treatment regimen, virological response, and adverse drug event (ADE) data were collected at the initial visit and 4, 8, 12, and 24 weeks later. The rapid virologic response (RVR) and sustained virologic response (SVR) rates were 90.4% and 97.8% for HCV genotype 1, 89.2% and 98.7% for genotype 6, 92.8% and 99% for genotype 2, and 90.9% and 100% for mixed genotype 2/6 or unspecified genotypes, respectively. There were no significant differences in the RVR and SVR rates for the various DAA regimens. The mean ALT, AST, and GGT activities decreased, and the PLT count increased during the treatments. ADEs occurred in 8% of the patients. The commonest ADEs were itching (3.1%), fatigue (1.8%), and dizziness (1.1%). None of the patients discontinued treatment because of an ADE. Posttreatment disease progression occurred in 7.7% of the patients, including liver fibrosis (3.6%), cirrhosis (1.1%), hepatocellular carcinoma (1.1%), and high alpha-fetoprotein (AFP) (1%). The factors associated with the achievement of RVR were low viral load, the use of sofosbuvir/ledipasvir or sofosbuvir/daclatasvir regimens, and a treatment duration of 12 weeks. No specific factors were found to be associated with the achievement of SVR. Posttreatment disease progression was associated with a high AFP and the use of sofosbuvir/ledipasvir. Thus, DAAs are highly effective and well-tolerated means of treating chronic HCV, and significantly improve patient outcomes. Their high efficacy and favorable safety profiles highlight the importance of early diagnosis and the use of personalized treatment strategies.

Molecular detection and genotyping of HBV from HBsAg positive patients in a tertiary hospital in Nigeria.

Nigeria remains one of the countries with a high hepatitis B virus (HBV) burden in Africa. Reports have indicated the presence of mixed HBV genotypes in Nigeria; however, there is still paucity of data regarding mixed genotype infections particularly in the Southern part of the country. Our aim is to determine the HBV genotype distribution among HBsAg-positive gastroenterology patients at the University College Hospital Ibadan, Nigeria. Serum samples were screened for HBsAg by ELISA, and positive samples were genotyped by semi-nested multiplex PCR for HBV genotypes A, B, C, D, E and F. Data generated were analyzed in R-studio. A total of 81/90 (90%) of HBsAg-positive samples were successfully genotyped, and genotype A was most prevalent with 15.7%, while genotypes B and E were the least with 1.2% each. Genotypes A/C infection was the highest among mixed infections with 40% prevalence, while genotypes A/D were the least prevalent mixed infection with 4.8%. We advocate for a comprehensive genotype analysis in larger cohorts across Nigeria, to give a more comprehensive understanding of the distribution and prevalence of different HBV genotypes population wide.

Does seasonal flowering and fruiting patterns of cacao only depend on climatic factors? The case study of mixed genotype populations in Côte d'Ivoire

Theobroma cacao, a tropical cauliflorous fruit tree, typically produces flowers and fruits twice a year and exhibits alternate harvesting patterns over consecutive six-monthly seasons in regions with bi-modal rainfall distribution, such as Côte d'Ivoire. This study investigated seasonal variations in flowering and fruiting among trees in populations of mixed cacao genotypes. The intensities of crown and trunk flowering and pod production were monitored for eight consecutive six-monthly seasons on 114 adult cacao trees grown from seedlings. We investigated distributions of seasonal flowering and fruiting values, relationships between flowering and pod production, and the effects of seasonal cumulative rainfall. The patterns of seasonal flowering and fruiting series were analyzed using two descriptors: the first distinguishing between regular and variable patterns, and the second analyzing the structure of such variability, classifying it as either irregular or alternating. Despite being subjected to similar climate and agronomic management, individual trees exhibited highly variable flowering and fruiting behaviors within each season, as well as variable patterns of flowering and fruiting across seasons. Seasonal alternate fruiting on a population scale masked highly variable patterns among trees, and only 19 % of the trees exhibited marked alternate fruiting patterns. Variations in pod production on a tree scale were mainly related to variations in trunk flowering. Endogenous factors seemed to control seasonal variations in flowering and fruiting, even though exogenous factors, both climatic and agronomic, could structure flowering and fruiting patterns at the orchard scale.

Intraspecies interactions of Streptococcus mutans impact biofilm architecture and virulence determinants in childhood dental caries.

Early childhood dental caries (ECC) is the most common chronic disease among children, especially among low socioeconomic populations. Streptococcus mutans is most frequently associated with initiation of ECC. Although many studies report children with multiple S. mutans strains (i.e., genotypes) have greater odds of developing ECC, studies investigating intraspecies interactions in dental caries are lacking. This study investigates the impact of intraspecies interactions on cariogenic and fitness traits of clinical S. mutans isolates using in vitro and in vivo approaches. Association analysis evaluated if presence of multiple S. mutans genotypes within the first year of colonization was associated with caries. Initially, clinical S. mutans isolates from 10 children were evaluated. S. mutans strains (G09 and G18, most prevalent) isolated from one child were used for subsequent analysis. Biofilm analysis was performed for single and mixed cultures to assess cariogenic traits, including biofilm biomass, intra-polysaccharide, pH, and glucan. Confocal laser scanning microscopy (CLSM) and time-lapse imaging were used to evaluate spatial and temporal biofilm dynamics, respectively. A Drosophila model was used to assess colonization in vivo. Results showed the mean biofilm pH was significantly lower in co-cultured biofilms versus monoculture. Doubling of S. mutans biofilms was observed by CLSM and in vivo colonization in Drosophila for co-cultured S. mutans. Individual strains occupied specific domains in co-culture and G09 contributed most to increased co-culture biofilm thickness and colonization in Drosophila. Biofilm formation and acid production displayed distinct signatures in time-lapsed experiments. This study illuminates that intraspecies interactions of S. mutans significantly impacts biofilm acidity, architecture, and colonization.IMPORTANCEThis study sheds light on the complex dynamics of a key contributor to early childhood dental caries (ECC) by exploring intraspecies interactions of different S. mutans strains and their impact on cariogenic traits. Utilizing clinical isolates from children with ECC, the research highlights significant differences in biofilm architecture and acid production in mixed versus single genotype cultures. The findings reveal that co-cultured S. mutans strains exhibit increased cell density and acidity, with individual strains occupying distinct domains. These insights, enhanced by use of time-lapsed confocal laser scanning microscopy and a Drosophila model, offer a deeper understanding of ECC pathogenesis and potential avenues for targeted interventions.

Molecular epidemiology of hepatitis B, hepatitis C, and HIV-1 co-infections in Ethiopia: Implications for disease burden and intervention strategies

BackgroundHepatitis B virus (HBV) exhibits high prevalence rates within Ethiopia. The genetic diversity of HBV, marked by mixed genotype infections, may hold significant implications for the trajectory of disease and responses to treatment. Ethiopia grapples with a substantial public health challenge posed by co-infections involving HBV, hepatitis C virus (HCV), and human immunodeficiency virus 1 (HIV-1), particularly among vulnerable populations. MethodsA comprehensive investigation into HBV, HCV, and HIV-1 co-infection was conducted. A total of 7,789 blood samples were meticulously analyzed, among which 815 exhibited HBV positivity. Among the HBV-positive samples, 630 were subjected to genotyping procedures, resulting in the identification of a prevalent trend of mixed infections characterized by HBV genotypes A/E/F (67.30%). Serological assessments were performed on 492 specimens to ascertain the presence of HCV and HIV-1 co-infections, revealing respective co-infection rates of 13.02% for HBV/HIV, 3.31% for HBV/HCV, and 2.07% for triple infection. ResultsThe investigation revealed the intricate prevalence of co-infections in Ethiopia, notably underlining the continued transmission of viruses. The prominent occurrence of mixed HBV genotypes A/E/F suggests dynamic viral interactions and ongoing transmission pathways. These findings accentuate the necessity for targeted interventions and enhanced patient care, as co-infections carry significant clinical complexities. ConclusionsThis study furnishes crucial insights into the molecular epidemiology of HBV, HCV, and HIV-1 co-infections in Ethiopia. The acquired knowledge can contribute to the advancement of strategies for clinical management and the formulation of public health interventions aimed at ameliorating the burden of viral infections within the nation.

Simultaneous genetic transformation and genome editing of mixed lines in soybean (Glycine max) and maize (Zea mays)

Robust genome editing technologies are becoming part of the crop breeding toolbox. Currently, genome editing is usually conducted either at a single locus, or multiple loci, in a variety at one time. Massively parallel genomics platforms, multifaceted genome editing capabilities, and flexible transformation systems enable targeted variation at nearly any locus, across the spectrum of genotypes within a species. We demonstrate here the simultaneous transformation and editing of many genotypes, by targeting mixed seed embryo explants with genome editing machinery, followed by re-identification through genotyping after plant regeneration. Transformation and Editing of Mixed Lines (TREDMIL) produced transformed individuals representing 101 of 104 (97%) mixed elite genotypes in soybean; and 22 of 40 (55%) and 9 of 36 (25%) mixed maize female and male elite inbred genotypes, respectively. Characterization of edited genotypes for the regenerated individuals identified over 800 distinct edits at the Determinate1 (Dt1) locus in samples from 101 soybean genotypes and 95 distinct Brown midrib3 (Bm3) edits in samples from 17 maize genotypes. These results illustrate how TREDMIL can help accelerate the development and deployment of customized crop varieties for future precision breeding.

Molecular Characterization of Hepatitis B Virus among HIV Positive and HIV Negative Pregnant Antenatal Women in Federal Capital Territory Abuja Hospital

A significant contributor to chronic hepatitis, cirrhosis and hepatocellular carcinoma is the Hepatitis B Virus (HBV). Hepatitis B Virus (HBV) is a major cause of Chronic Hepatitis, cirrhosis and Hepatocellular Cancer (HCC). The incidence of HBV-related HCC cases is projected to increase for at least two decades due to the high prevalence of chronic HBV infection throughout the world. The overall aim of this study is to determine the sero-prevalence and molecular characterization of hepatitis B virus among HIV positive and HIV negative antenatal women attending Federal Capital Territory Abuja hospitals. A total number of 302 samples were collected from 302 participants between the ages of 18-55 years. The samples were screened for HIV, HBV, HBV serology markers, using rapid test kits, HBV Viral load and HBV genotyping was carried out using multiplex PCR method and data was analyzed using appropriate tools. Nine tested positive for HIV, 34 tested positive for HBV and 2 tested positive for both HIV and HBV, corresponding to 2.9 % (95% CI; 1.2 – 3.8) prevalence for HIV, 11.3% (95% CI; 8.7 – 13.6) for Hepatitis B and 0.7% for HIV/HBV co-infection. Age 25-35 years old had the highest prevalence of HBV 22(7.3%) followed by those of less than 25 years old 8(2.8%) and 1(0.7%) each for age range 36-45 years, 2(0.7%) and 46-55 years. HBeAb and HBcAb were expressed highest among subject within the age range of 25 – 35 year 14 (41.2%) and 22 (64.7 %,) participants respectively, followed by the age range of less than 25 years 5 (14.7%) and 8 (36.4%), for HBeAb and HBcAb respectively. There was no Hepatitis B envelop antigen (HBeAg) nor Hepatitis B surface antibody (HBsAb) among the study group. Of the 34 samples, 20 (62%) had undetectable viral load, while 14 (38%) had detectable viral load. Genotype E was three times more prevalent among those of 25 years old and above than those less than 25 years old detected (75% and 25%). However, genotype B/E mixed was more expressed (60% of cases) among those less than 25 years old, than those greater than 25 years of age (40%). There was no significant difference in the prevalence of genotype E and B/E serotype between HBV+/HIV-patients and their HBV+/HIV+ counterpart (P=0.08 and P=0.15, respectively). HBV Genotype E single infection viral load was significantly (P=0.01) more expressed among those 25 years of age and above than less than 25 years old (4,354,703 ± 346865 IU/ml versus 55.6 ±28.1 IU/ml). Mixed infection genotype B/E was significantly (P=0.04) more expressed among those less than 25 years of age than those 25 years old and above (730.9 ± 238 IU/ml versus IU/ml versus 91.1 ± 48 IU/ml). There was no significant difference between the mean of HBV Viral load HBV+/HIV- patients and their HBV+/HIV+ counterpart (T-test =0.009; P=0.92). In conclusion FCT is 11.3% prevalence rate of HBV and genotypes E and B/E are in circulation among pregnant women among the pregnant women attending antenatal clinic in FCT hospital in Abuja.

Investigating Dirofilaria immitis isolates infecting domestic canines and their susceptibility/resistance patterns to macrocyclic lactones in the northern region of the Mississippi Delta area (southeast Missouri)

Previous reports of macrocyclic lactone (ML) resistance in Dirofilaria immitis, the parasitic nematode which causes heartworm disease, have mainly been from the southern Mississippi Delta region. Southeast Missouri (SEMO), forming the northern boundary of this region, has not previously been well studied. The area is an ideal propagation region for heartworm infection and possibly for the spread of ML resistance. To assess whether D. immitis isolates infecting domestic canines in SEMO exhibit evidence of resistance to MLs, domestic canines, presented to veterinary facilities testing positive for heartworms through antigen and microfilariae (MF) examination, were utilized in the study. Using a descriptive epidemiological cross-sectional study, from March 2021 through February 2022, blood sample collection from 96 canines living in SEMO testing positive for heartworms were analyzed. MiSeq technology was utilized to sequence specific genetic markers associated with susceptibility/resistance for MLs in D. immitis isolates. Genomic data revealed most D. immitis isolates had genotypic profiles consistent with resistance to MLs. Of the 96 samples tested, 91 (94.8%) had a resistant genotype, 4 (4.2%) had a mixed genotype, and 1 sample (1%) genotyped as susceptible. While detailed and reliable medical histories were not available for most canines, detailed medical history from 2 canines indicated evidence of phenotypic resistance that was consistent with their genotypes. However, in vivo preventive tests are needed to confirm a high frequency of phenotypic ML resistance in D. immitis from this region. Increasing resistance patterns to MLs indicate the approach to heartworm prevention/treatment protocol should be reconsidered. New measures may be required to stop heartworm disease.

Sub-microscopic Plasmodium falciparum infections and multiple drug resistant single nucleotide polymorphic alleles in pregnant women from southwestern Nigeria

ObjectivesThe study evaluated sub-microscopic malaria infections in pregnancy using two malaria Rapid Diagnostic Tests (mRDTs), microscopy and RT-PCR and characterized Plasmodium falciparum dihydrofolate reductase (Pfdhfr) and Plasmodium falciparum dihydropteroate synthase (Pfdhps) drug resistant markers in positive samples.MethodsThis was a cross sectional survey of 121 pregnant women. Participants were finger pricked, blood drops were collected for rapid diagnosis with P. falciparum histidine-rich protein 11 rapid diagnostic test kit and the ultra-sensitive Alere Pf malaria RDT, Blood smears for microscopy and dried blood spots on Whatman filter paper for molecular analysis were made. Real time PCR targeting the var acidic terminal sequence (varATS) gene of P. falciparum was carried out on a CFX 96 real time system thermocycler (BioRad) in discriminating malaria infections. For each run, laboratory strain of P. falciparum 3D7 and nuclease free water were used as positive and negative controls respectively. Additionally, High resolution melt analyses was employed for genotyping of the different drug resistance markers.ResultsOut of one hundred and twenty-one pregnant women sampled, the SD Bioline™ Malaria Ag P.f HRP2-based malaria rapid diagnostic test (mRDT) detected eight (0.06%) cases, the ultra-sensitive Alere™ malaria Ag P.f rapid diagnostic test mRDT had similar outcome in the same samples as detected by the HRP2-based mRDT. Microscopy and RT-PCR confirmed four out of the eight infections detected by both rapid diagnostic tests as true positive and RT-PCR further detected three false negative samples by the two mRDTs providing a sub-microscopic malaria prevalence of 3.3%. Single nucleotide polymorphism in Pfdhps gene associated with sulphadoxine resistance revealed the presence of S613 mutant genotypes in three of the seven positive isolates and isolates with mixed wild/mutant genotype at codon A613S. Furthermore, four mixed genotypes at the A581G codon were also recorded while the other Pfdhps codons (A436G, A437G and K540E) showed the presence of wild type alleles. In the Pfdhfr gene, there were mutations in 28.6%, 28.6%, and 85.7% at the I51, R59 and N108 codons respectively. Mixed wild and mutant type genotypes were also observed in 28.6% each of the N51I, and C59R codons. For the Pfcrt, two haplotypes CVMNK and CVIET were observed. The SVMNT was altogether absent. Triple mutant CVIET 1(14.3%) and triple mutant + wild genotype CVIET + CVMNK 1(14.3%) were observed. The Pfmdr1 haplotypes were single mutants YYND 1(14.3%); NFND 1(14.3%) and double mutants YFND 4(57.1%); YYDD 1(14.3%).

Factors associated with keel bone damage – a longitudinal study of commercial layer flocks during the laying period

ABSTRACT 1. Keel bone damage, such as deformations and fractures, is a severe problem regarding animal welfare in layers. To identify risk factors under commercial conditions, 33 layer flocks (22 barn, 11 free range) with white (n = 18), brown (n = 11) and mixed (n = 4) genotypes were examined. 2. Keel bone status was frequently scored by palpation throughout the laying period. Data on housing and management conditions were collected. Multiple regression and Generalized Estimating Equations procedure were used for analysis. 3. At 65–74 weeks of age, the prevalence of keel bone damage ranged between 26% and 74%. White genotypes and those kept in multi-tier systems developed significantly (p < 0.05) more keel bone damage than brown genotypes or those kept in single-tier systems. Wing feather condition was associated with keel bone damage (p < 0.05), while other investigated variables regarding health, housing and management were not associated. 4. In conclusion, housing and management should be adapted to meet the birds’ specific needs in multi-tier systems, which may vary for brown and white genotypes. Whether those differences result from genotype associated predispositions or other individual traits remains to be determined.

Chd8 haploinsufficiency impacts rearing experience in C57BL/6 mice.

Mutations in CHD8 are one of the highest genetic risk factors for autism spectrum disorder. Studies in mice that investigate underlying mechanisms have shown Chd8 haploinsufficient mice display some trait disruptions that mimic clinical phenotypes, although inconsistencies have been reported in some traits across different models on the same strain background. One source of variation across studies may be the impact of Chd8 haploinsufficiency on maternal-offspring interactions. While differences in maternal care as a function of Chd8 genotype have not been studied directly, a previous study showed that pup survival was reduced when reared by Chd8 heterozygous dams compared with wild-type (WT) dams, suggesting altered maternal care as a function of Chd8 genotype. Through systematic observation of the C57BL/6 strain, we first determined the impact of Chd8 haploinsufficiency in the offspring on WT maternal care frequencies across preweaning development. We next determined the impact of maternal Chd8 haploinsufficiency on pup care. Compared with litters with all WT offspring, WT dams exhibited less frequent maternal behaviors toward litters consisting of offspring with mixed Chd8 genotypes, particularly during postnatal week 1. Dam Chd8 haploinsufficiency decreased litter survival and increased active maternal care also during postnatal week 1. Determining the impact of Chd8 haploinsufficiency on early life experiences provides an important foundation for interpreting offspring outcomes and determining mechanisms that underlie heterogeneous phenotypes.

Genomic and common garden data reveal significant genetic differentiation in the endangered San Fernando Valley spineflower Chorizanthe parryi var. fernandina

San Fernando Valley spineflower (Chorizanthe parryi var. fernandina [S. Watson] Jeps.) (Polygonaceae) is an herbaceous annual plant, endemic to California, and until rediscovered in 1999 had been thought to be extinct for almost seven decades. Historically documented at 10 locations, it currently persists at 2, separated by approximately 27 km. State listed as endangered, a description of its genetic diversity and structure is of conservation interest. After determining a lack of variation in ploidy, we examined genetic variation from samples within both populations: a common garden study for potentially adaptive genetic variation in selected growth and phenological traits and analysis of single nucleotide polymorphisms identified through restriction-site associated DNA sequencing. Both measures indicated that this highly restricted taxon nevertheless harbors substantial levels of genetic diversity and has significant between- and within-population genetic structure. Combining approaches from population genomics and common garden studies provided more insight into the patterns and basis of genetic diversity than is typical for studies of non-model species. Although local adaptation was not specifically studied (i.e., via reciprocal transplant studies), the differences determined from these two independent lines of evidence indicate that mixing gene pools between populations is not recommended at this time. Further, with significant differences revealed among subpopulations, we caution against mixing genotypes across subpopulations for the most part, and without much more evidence that this would not pose a risk of outbreeding depression. The importance of supporting pollinator health and diversity is highlighted. With genetic diversity—particularly with an annual species—being dynamic, fluctuating with the usual processes and with contributions from the soil seedbank, we recommend periodic resampling to monitor genetic diversity and structure. Climate change is anticipated to contribute to this variability.

Brown rats (Rattus norvegicus) as potential reservoirs of Enterocytozoon bieneusi in Heilongjiang Province, China: high prevalence, genetic heterogeneity, and potential risk for zoonotic transmission.

Enterocytozoon bieneusi, an obligatory intracellular fungus, is prevalent among animals and humans. Due to their close interaction with humans and their extensive regional distribution, brown rats (Rattus norvegicus) are important pathogen reservoirs. To assess the zoonotic transmission potential of E. bieneusi, a molecular investigation was conducted on 817 R. norvegicus from four cities in Heilongjiang Province, China. A total of 817 R. norvegicus were collected from four cities in Heilongjiang Province, China. The genotyping of E. bieneusi was conducted through PCR amplification of the small subunit ribosomal RNA (SSU rRNA)'s internal transcribed spacer (ITS) segments. Phylogenetic and similarity analyses were used to examine zoonotic potential and genetic characteristics of the E. bieneusi-positive specimens. Among the 817 R. norvegicus, the total infection rate was 33.3% (272/817). Seventy-five genotypes were identified, including 14 known genotypes D (n = 167), A (n = 15), HLJ-CP1 (n = 12), WR8 (n = 6), EbpC (n = 2), BEB6 (n = 1), CS-4 (n = 1), CHPM1 (n = 1), Henan-II (n = 1), HNH-22 (n = 1), HNH-25 (n = 1), I (n = 1), JLD-XI (n = 1), SDD5 (n = 1), and 61 novel genotypes designated as SHWR1 (n = 10), SYSWR1 (n = 2), and SHWR2 to SHWR17, SYSWR2 to SYSWR36 and QTHWR1 to QTHWR8 (n = 1, each). Moreover, 10 samples exhibited mixed genotype infections, including D + A (n = 3), D + EbpC (n = 1), D + HLJ-CP1 (n = 1), D + SHWR1 (n = 1), D + SHWR16 (n = 1), D + SHWR17 (n = 1), SDD5 + WR8 (n = 1), and CS-4 + SYSWR36 (n = 1). Phylogenetic analysis grouped the genotypes into three main groups: group 1 (n = 67), group 2 (n = 5), and group 9 (n = 3). The high prevalence and genetic diversity of E. bieneusi in Heilongjiang Province's R. norvegicus imply that these animals spread the pathogen. The R. norvegicus that E. bieneusi carries can spread zoonotic disease, making it a serious hazard to the local human population. Therefore, it is imperative to raise awareness about the dangers posed by R. norvegicus and implement measures to reduce their population to prevent environmental contamination.

904. Epidemiology of Rotavirus Group A Mixed Genotypes Isolated from Children with Acute Gastroenteritis over a 15-Year Period

Abstract Background Limited data exist worldwide regarding the genetic epidemiology of mixed G and/or P Rotavirus group A (RVA) strains and their association with demographic and clinical characteristics in children with acute gastroenteritis (AGE). Methods Positive fecal samples for mixed RVA previously G (VP7) and P (VP4) genotyped from children aged ≤16 years with AGE between 2007-2021 (15 years) in Greece were further genotyped for I (VP6) and E (NSP4) using RT-PCR and Sanger sequencing. Demographic, clinical and laboratory data of children infected with a mixed RVA strain were also collected. Results Over the 15-year period, 129/4427 (2.9%) mixed RVA strains were detected. Higher detection rate was observed in 2016 (7.5%) and in 2021 (5.8%) (Figure 1). Children infected with a mixed RVA strain had a median age of 21.1 months (IQR: 56.2) and were predominantly males (85/129, 65.9%). Mixed G genotype were 73/129 (56.6%), mixed P type 54/129 (41.1%) and both G and P mixed genotypes 3/129 (2.3%). In total, 27 different G/P combinations were detected with most prevalent the: G1G4P[8] (47/129, 36.4%), G1P[4]P[8] (18/129, 14%) and G4P[4]P[8] (18/129, 14%) (Figure 2). Higher seasonal peaks were observed for G mixed types in spring (68.8%) and for P mixed types in summer (66.7%). Infection with a G mixed type had a higher relative frequency in children aged 0-12 months, while infection with P mixed type had a higher relative frequency in children aged 13-48 months (p=0.043). I and E typing was successful in 49.6% (64/129) of mixed samples from 2013-2021. Two different I (I1, I2) and three different E (E1, E2, E3) genotypes were detected, with I1 (42.2%, 27/64) and E1 (39.1%, 25/64) being the most common I and E genotypes, respectively. In 31.3% (20/64) of samples, mixed I (I1I1, I1I2) and/or E (E1E1, E1E2) genotypes were also detected. No association has been detected between mixed genotypes and clinical characteristics of the patients. Annual distribution of total and mixed RVA cases in a 15-year period G-P combinations of RVA mixed strains. Other combinations found in a smaller proportion (&amp;lt;2.3%) were: G3P[4]P[8] (1.6%), G4P[8]P[9] (1.6%), G6P[4]P[9] (1.6%), GUDP[4]P[8] (1.6%), G2G4P[4]P[8] (1.6%), G1G2P[4] (0.8 %), G1G3P[9] (0.8%), G1G8P[4]P[8] (0.8%), G10P[8]P[10] (0.8%), G12P[4]P[8] (0.8%), G2P[8]P[10] (0.8%), G2G4P[8] (0.8%), G2G9P[8] (0.8%), G4G12P[4] (0.8%), G8G9P[8] (0.8%), G9P[4]P[8] (0.8%), G9P[6]P[8] (0.8%), G9G12P[8](0.8%). Conclusion These results indicate the great genetic diversity of RVA strains. Significant differences were found in the seasonality of the mixed G and P strains as well as in the age group of children infected with mixed G and P strains. Disclosures All Authors: No reported disclosures

Molecular evolution, virology and spatial distribution of HCV genotypes in Pakistan: A meta-analysis

BackgroundHepatitis C, caused by the Hepatitis C Virus (HCV), is the second most common form of viral hepatitis. The geographical distribution of HCV genotypes can be quite complex, making it challenging to ascertain the most prevalent genotype in a specific area. MethodsTo address this, a review was conducted to determine the prevalence of HCV genotypes across various provinces and as a whole in Pakistan. The scientific literature regarding the prevalence, distribution, genotyping, and epidemiology of HCV was gathered from published articles spanning the years 1996–2020. ResultsGenotype 1 accounted for 5.1% of the patients, with its predominant subtype being 1a at 4.38%. The frequencies of its other subtypes, 1b and 1c, were observed to be 1.0% and 0.31% respectively. Genotype 2 had a frequency of 2.66%, with the most widely distributed subtype being 2a at 2.11% of the patients. Its other subtypes, 2b and 2c, had frequencies of 0.17% and 0.36% respectively. The most prevalent genotype among all isolates was 3 (65.35%), with the most frequent subtype being 3a (55.15%), followed by 3b (7.18%). The prevalence of genotypes 4, 5, and 6 were scarce in Pakistan, with frequencies of 0.97%, 0.08%, and 0.32% respectively. The prevalence of untypeable and mixed genotypes was 21.34% and 3.53% respectively. Estimating genotypes proves to be a productive method in assisting with the duration and selection of antiviral treatment. Different HCV genotypes can exhibit variations in their response to specific antiviral treatments. Different genotypes may have distinct natural histories, including variations in disease progression and severity. Some genotypes may lead to more rapid liver damage, while others progress more slowly. ConclusionsThis information can guide screening and testing strategies, helping to identify individuals at higher risk of developing severe complications. Studying the distribution of HCV genotypes in a population can provide valuable insights into the transmission dynamics of the virus.

Resilience of Emiliania huxleyi to future changes in subantarctic waters.

Lower pH and elevated temperature alter phytoplankton growth and biomass in short-term incubations, but longer-term responses and adaptation potential are less well-studied. To determine the future of the coccolithophore Emiliania huxleyi, a mixed genotype culture from subantarctic water was incubated for 720 days under present-day temperature and pH, and also projected future conditions by the year 2100. The future population exhibited a higher growth rate relative to present-day cells transferred to future conditions after 309 days, indicating adaptation or genotype selection; this was reflected by an increase in optimum growth temperature of ~2.5°C by the end of the experiment. Following transfer to opposing conditions in short-term cross-over incubations, cell volume responded rapidly, within eight generations, confirming trait plasticity. The changes in growth rate and cell volume were larger than reported in previous single stressor relationships and incubations, suggesting synergistic or additive effects of combined elevated temperature and lower pH and highlighting the importance of long-term multiple stressor experiments. At the end of the incubation there were no significant differences in cellular composition (particulate organic content and chlorophyll a), or primary production between present-day and future populations. Conversely, two independent methods showed a 50% decrease in both particulate inorganic carbon and calcification rate, consistent with the decrease in cell volume, in the future population. The observed plasticity and adaptive capacity of E. huxleyi indicate resilience to future conditions in subantarctic waters, although changes in cell volume and carbonate may alter grazing loss and cell ballast, so influencing carbon export to the deep ocean.

Universal primer multiplex PCR assay for detection and genotyping of porcine astroviruses

Porcine astroviruses (PAstV) are members of the family Astroviridae, Mamastravirus genus and have been identified to have five genotypes (PAstV1–5). These viruses are highly prevalent in pigs and can cause enteric disease as well as neurological or respiratory symptoms depending on their genotypes. At present, the epidemiological impacts of some PAstV genotypes on pigs are largely unknown and hence continuously monitoring of these PAstVs may be needed. The purpose of this research was to develop an improved and efficient detection tool for PAstVs and to evaluate the developed method using clinical samples. Initially, a set of five chimeric primers (CP), each comprising genotype specific primer pairs with an identical universal adapter at the 5’ end, and a universal primer (UP) that is identical to universal adapter sequence, were designed. With these tools in place, a novel multiplex PCR system with universal primer was established for the simultaneous detection of the five types of PAstV. This method can specifically detect PAstV genotypes, with a limit of detection (LOD) of 5 copies/μL for each genotype irrespective of single or mixed target template. Using this new assay, 273 pig fecal samples were investigated for further assay evaluation. Among all samples, the positive rate was 70.0% with PAstV4 in 56.8% of the samples, PAstV2 in 38.8%, PAstV1 in 16.8%, and PAstV5 in 11.0%. More than one PAstV in a sample were detected in 39.2% of the samples. The consistency rate between the novel multiplex PCR and singleplex PCRs was 96.4–100%. Given its rapidity, specificity and sensitivity, the novel multiplex PCR is a useful approach for demonstrating single or mixed genotype infections of PAstV.

A Retrospective Study to Determine the Genotypic Distribution of Hepatitis-C from a Tertiary Care Hospital in South India

Hepatitis C virus (HCV) is a blood-borne pathogen that transmits infection via transfusion. Hepatocellular carcinoma is the fifth most common cancer and a major cause of death in patients with chronic HCV infection. Response to treatment is mainly based on the genotypic characterization of HCV. The gold standard for genotyping HCV is by sequencing highly conserved regions such as NS5, core, E1, and 5’UTR. Serum samples of patients who visited the tertiary care hospital with clinical features suggestive of HCV infection formed the study group. HCV genotyping was performed using multiplex Polymerase Chain Reaction in the samples tested positive by Chemiluminescence Immunoassay (CLIA). The viral loads were also performed on selected patient samples. In the present study, Genotype 4 (35.71%), followed by Genotype 3 (17.53%) and 1 &amp; 1b (12.34%) were the common genotypes observed. Genotype 1,1b &amp; 4 mixed type and genotype 4 and 5 mixed type was detected in one sample each (0.65%). The mean measured value of HCV antibody was 11.51 ± 4.57. The viral load was detected in 61 out of 81 samples tested. The mean viral load ranged from 550 to 552769250IU/ml (log 2.74-log 8.74). Genotype 4 was the most common genotype demonstrated in our study as opposed to the other studies were genotype 3 was the dominant one in south India.